Endocytic-autophagic-lysosomal system (EALS) abnormalities are invariant features of vulnerable neuronal populations in Alzheimer's disease (AD). Our studies have shown that abnormalities in early endosomes coincide with the earliest elevations in soluble Abeta and precede amyloid deposition in sporadic AD and Trisomy 21 (Ts21, Down syndrome) brain. Ts65Dn mice, a model of human Ts21, and Ts21 fibroblasts also display endosomal pathway pathologies. Our preliminary findings show that endocytic pathology extends beyond the early endosome to the late endosome - a compartment shared by both the endocytic and degradative pathways. Using various models, in Aim 1 and in conjunction with the other Projects of this Program, we will test our hypothesis that late endocytic abnormalities are a unifying feature of the EALS dysfunction seen in sporadic AD and Ts21, as well as in presenilin-linked familial AD with autophagic pathology. In Aim 2, we will confirm that structural changes in the late endocytic pathway result from altered functional relationships between compartments of the endocytic and autophagic pathways by following in vitro endocytic uptake and degradation; further, we will attempt to rescue pathogenic endocytic events in the Ts21 background by altering expression of key vesicular trafficking proteins (rab GTPases). Endocytic alterations are dependent upon App gene triplication in the trisomy background. In Aim 3, we will investigate the role that APP expression levels play in modulating endocytic alterations in the Ts65Dn background and examine the role of endocytic dysfunction in mediating the neurodegeneration seen in the Ts65Dn mouse. Additionally, using Ts65Dn crosses with APP overexpressing, APP knockout, BACE knockout and PS1 hypomorphic mice, we will examine whether betaC-terminal fragments or other parts of APP, Abeta, or APP itself modulate endocytic dysfunction and neurodegeneration. Crosses between Ts65Dn mice and an amyloid depositing APP overexpressing mouse will be used to test our hypothesis that the endocytic alterations seen in Ts21 and AD promote amyloidogenesis. Lastly, in Aim 4, we will use Ts21 models to determine if cholesterol and APOE4, important risk factors for AD, promote endocytic pathway abnormalities and, in Ts65Dn mice, lead to accelerated neurodegeneration and amyloidogenic APP processing.